The goal of this project is to definitively establish the role of the cell surface receptor TREM-1 in the Dathogenesis of sepsis. We hypothesize that activation of TREM-1 synergizes with ligation of the Toll-Like Receptors (TLRs) and amplifies the inflammatory response during sepsis. We will test this hypothesis with three specific aims. In the first specific aim, we will use a newly generated strain of mice in which we have humanized the TREM locus by deleting the murine TREM-3 gene (which is a pseudogene in humans) in tandem with the TREM-1 gene. This mouse is the ideal reagent to model the role of TREM-1 in human biology. Our preliminary data indicate that TREM-1/3 double deficient mice are more resistant to sepsis than wild type mice. We will rigorously test the response of this strain of mice in clinically relevant models of sepsis including cecal ligation and puncture and both gram-positive and gram-negative septic pneumonia. In the second specific aim, we will define the signaling pathway through which TREM-1 and the TLRs interact with a focus on identifying nodes in signaling cascades that could serve as therapeutic targets. The third specific aim is to identify the endogenous ligand for TREM-1. In our preliminary experiments we have established a system that detects a ligand on the surface of granulocytes of septic patients and have identified a monoclonal antibody that recognizes the TREM-1 ligand or a component of it. The ligand for TREM-1 represents a novel target for sepsis therapy. Our studies on TREM biology and our previous experience in the study of sepsis positions us well to succeed at our specific aims. We originally discovered and cloned the TREM genes and thus have an extensive collection of molecular-biological tools for manipulating the TREM system. We have successfully generated TREM-1/TREM-3 knockout animals to best model the role of TREM-1 in human biology. Importantly, we have well established collaborations with a multi-disciplinary group of investigators including active critical care physicians who can provide access to clinical materials and direction as to the clinical relevance of our data, experienced sepsis researchers who can provide expertise in animal models of sepsis, and an expert biochemist to collaborate in the identification of novel mediators. Taken together, we feel that this proposal will advance the study of sepsis by defining the biology of TREM-1 in sepsis, establishing the rnechanism by which TREM-1 amplifies inflammation and determining the ligand for TREM-1, a potential novel mediator of sepsis. In lay terms, the goal of this project is to define the role of the TREM-1 protein in the biology of sepsis, a leading cause of death in critically i patients. Our proposed experiments will define the function of TREM-1 using mouse studies that model the human disease and biochemical studies to identify the mechanisms by which TREM-1 functions in the cell. We also have detected a binding partner of TREM-1 (the TREM-1 ligand) and we propose to identify this molecule as it may serve as a novel target for the treatment of septic patients,